Itch is an unpleasant sensation associated with the desire to scratch. While acute itch commonly occurs with insect bites, allergic reactions or contact with certain plants, chronic itch is a common symptom of many dermatological conditions and systemic disease. Chronic itch is poorly-treated, negatively impacts the quality of life, and carries a heavy economic and social burden. There is a great need to better understand mechanisms of itch to develop more effective treatments. This proposal will use behavioral assessment of scratching, combined with electrophysiological and neuroanatomical approaches, to investigate mechanisms of itch transmission in normal animals and in animal models of chronic itch. The project consists of 5 Specific Aims. Specific Aim 1 will investigate if intradermal (id) microinjection of the itch mediators serotonin, agonists of protease-activated receptor (PAR) subtypes 2 and 4, and histamine, elicit scratching behavior in mice in a manner that is consistent with itch sensation in humans. We will test if scratching is reduced by -opioid antagonists but not by morphine, which often evokes itch. Specific Aim 2 seeks to identify the locations and functional properties of pruritogen-activated neurons in the superficial dorsal horn of the mouse. This will be accomplished using a neuroanatomical approach to localize pruritogen-sensitive neurons, and an electrophysiological approach to record the responses of neurons in these locations to pruritic and noxious stimulation of the skin in anesthetized mice. We hypothesize that a sub-population of neurons in superficial laminae of the murine lumbar dorsal horn responds to id pruritogens in a manner consistent with a role in signaling acute itch. Specific Aim 3 will investigate the role of the spinothalamic and spinoparabrachial pathways in mediating itch in mice and rats using a neuroanatomical double-label strategy. Specific Aim 4 will investigate a role for substance P and other neuropeptides in spinal itch transmission, using neurotoxic ablation of neurons expressing the NK-1 receptor as well as intracisternal delivery of antagonists. Specific Aim 5 will investigate sensitization of itch-signaling pathways in mouse models of chronic itch. We hypothesize that experimental chronic itch will lead to enhanced scratching elicited by pruritogens and also algogens such as capsaicin. We further hypothesize that experimental chronic itch sensitizes superficial dorsal horn neurons, manifested by increased Fos expression and functional hyperexcitability as exhibited by abnormal spontaneous activity and increased responses to pruritogens. These hypotheses will be addressed using two murine models of chronic itch, the NC itchy mouse strain, and experimental dry skin.